The long-term goal of this study is to understand how photoresponses are forwarded through the vertebrate retina to higher visual centers. This proposal is aimed at determining the molecular identity of the voltagegated calcium channel pore-forming alphal subunit isoforms underlying the L-type calcium currents of photoreceptors and bipolar ceils of primate retina. Immunolabeling of different alpha-1 subunit isoforms combined with immunolabeling of cone opsins will test the hypothesis that expression of specific isoforms is restricted to spectral classes of cones. Antibody studies will be combined with isoform-specific RT-PCR amplification and sequencing of transcripts to determine molecular identity of expressed alphal subunit isoforms. Studies will be performed on whole retina and eventually on single, isolated cells to establish the diversity of expression at the single cell level. To determine the role of the different isoforms in the regulation of the intracellular calcium dynamics of the soma and synaptic terminals of these cells, electrophysiology combined with optical imaging of calcium indicators will be applied. Selective blockage of calcium entry at restricted sites, or controlled ablation of cellular compartments, will indicate the location of expression of calcium channels on cones and bipolar cells. These studies will provide the foundation for further studies aimed at understanding the mechanisms that control the regional expression of calcium channel proteins and their relationship to spatially-restricted cellular processes such as transmitter release. Another aim of this study is to identify novel candidate genes for human retinal channelopathies involving abnormal processing of photoreceptor and/or bipolar cells. In humans, mutations in the alphalF calcium channel subunit gene, CACNA1F, have been linked to one form of night blindness (CSNB2), underscoring the importance of this calcium channel subunit in normal visual processing. Newly identified calcium channel alphal subunit isoforms will become candidate genes in human visual disorders involving photoreceptor and/or bipolar cell dysfunction. This study will therefore add to our current understanding of blindness as well as our fundamental understanding of the structure and function of retinal neurons.